Involvement of myosin light-chain kinase in endothelial cell retraction.
Permeabilized bovine pulmonary artery endothelial cell monolayers were used to investigate the mechanism of endothelial cell retraction. Postconfluent endothelial cells permeabilized with saponin retracted upon exposure to ATP and Ca2+. Retraction was accompanied by thiophosphorylation of 19,000-Da myosin light chains when adenosine 5'-[gamma-[35S]thio]triphosphate was included in the medium. Both retraction and thiophosphorylation of myosin light chains exhibited a graded quantitative dependence on Ca2+. When permeabilized monolayers were extracted in buffer D containing 100 mM KCl and 30 mM MgCl2 for 30 min, the cells failed to retract upon exposure to ATP and Ca2+, and no thiophosphorylation of myosin light chains occurred. The ability both to retract and to thiophosphorylate myosin light chains was restored by the addition to the permeabilized, extracted cells of myosin light-chain kinase and calmodulin together but not by either alone. These studies indicate that endothelial cell retraction, as does smooth muscle contraction, depends on myosin light-chain kinase phosphorylation of myosin light chains.
Wysolmerski
R B
RB
Department of Pathology, Saint Louis University School of Medicine, MO 63104.
Lagunoff
D
D
D000199:Actins / Q000032:analysis; D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism; D000818:Animals; D002417:Cattle; D002460:Cell Line; D002463:Cell Membrane Permeability; D004591:Electrophoresis, Polyacrylamide Gel; D004730:Endothelium, Vascular / Q000502:physiology*; D008856:Microscopy, Fluorescence; D008970:Molecular Weight; D015879:Myosin Subfragments / Q000302:isolation & purification / Q000378:metabolism; D009219:Myosin-Light-Chain Kinase / Q000302:isolation & purification / Q000378:metabolism*; D013462:Sulfur Radioisotopes
D000199:Actins; D015879:Myosin Subfragments; D013462:Sulfur Radioisotopes; C022571:adenosine 5'-O-(3-thiotriphosphate); D000255:Adenosine Triphosphate; D009219:Myosin-Light-Chain Kinase